Gene therapy for the thalessemia syndromes requires that the introduced globin gene mimic the function of an in situ normal globin gene both in tissue specificity and efficiency of transcription. DNA mediated gene transfer experiments have shown that the human beta-globin gene with its immediate 5' and 3' flanking sequences can be expressed in a tissue and developmental-stage specific manner. Yet high level transcription of such a gene may require sequence elements not residing in the globin structural gene and its immediate flanking sequences. Comparison of the extents of DNA deletions in the Dutch and English gamma delta beta-thalassemias suggests the possible existence of regulatory sequences upstream of the embryonic epsilon-globin gene, which can regulate the expression of the far downstream beta-globin gene. A segment of DNA at 10 Kb 5' of the epsilon-globin gene has been shown to display erythroid specific enhancer activity. It is therefore possible that transcriptional activation of the beta-globin gene may involve at least two synergistic activation steps, mediated respectively by this distant enhancer sequence, and by sequences much closer to the beta-globin gene. In DNA mediated gene transfer experiments, the presence of this enhancer element, in cis to the beta-globin or a test gene, may thus significantly enhance the transcriptional efficiency of such a gene in erythroid hosts. This proposal attempts to address this possibility. The enhancer element with other probable regulatory sequences will be spliced with a beta-globin or a test gene into enhancerless plasmids or into an enhancerless retroviral vector, which will subsequently be transiently or stably introduced into appropriate cell lines or into mouse bone marrow cells. The transcriptional efficiency of the test gene will be compared to that of the endogenous alpha-or beta-globin gene of the erythroid host cells. Identification of an erythroid specific, globin gene enhancer, understanding the molecular mechanism of its action on the beta- like globin genes, and its possible role in the coordinated activation of the alpha- and beta-like globin genes located on separate chromosomes, may all aid in elucidating the regulatory mechanism of the human beta-globin gene and thus in bringing gene therapy for the beta-thalassemias significant steps closer to reality.